Unveiling the dominant peptides recognized by chicken TCRγδ+CD8α+ T cells against avian leukosis virus subgroup J (ALV-J) infection in B21 haplotype chickens

doi:10.1016/j.jia.2026.07.010

Journal of Integrative Agriculture

2026, Vol. 25

Issue (10): 3990-4001 DOI: 10.1016/j.jia.2026.07.010

Special Focus: Advancing China–Africa Agricultural Science and Technology Cooperation: Innovation and Pathways

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Unveiling the dominant peptides recognized by chicken TCRγδ⁺CD8α⁺ T cells against avian leukosis virus subgroup J (ALV-J) infection in B21 haplotype chickens

Xueqing Li¹, Tianqi Teng¹, Yingyi Chen¹, Zhuohang Li¹, Jiangwu Huang¹, Tao Zhang¹, Irene Ogali³, Manman Dai^{1, 2#}

¹Guangdong Laboratory for Lingnan Modern Agriculture, National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China

²UK-China Centre of Excellence for Research on Avian Diseases, Guangzhou 510642, China

³Veterinary Science Research Institute–Kenya Agricultural and Livestock Research Organization, P.O.Box 32-00902 Kikuyu, Kenya

Highlights
● ALV-J infection induces the expansion and activation of TCRγδ⁺CD8α⁺T cells in vivo and in vitro.
● TCRγδ⁺CD8α⁺ T cell responses to ALV-J infection are more predominant in H-B21 chickens than in G-WL chickens.
● Gag_187–215 and Pol_417–445 are dominant ALV-J peptides recognized by chicken TCRγδ^⁺CD8α⁺ T cells.

Abstract
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摘要

TCRγδ⁺T细胞是鸡体内重要的淋巴细胞群体，但其对J亚群禽白血病病毒（Avian leukosis virus subgroup J，ALV-J）的应答特征及其所识别的抗原仍缺乏深入研究。本研究发现，ALV-J感染后，B21单倍型鸡（H-B21）和不携带MHC B21的SPF鸡（G-WL）体内TCRγδ⁺CD8α⁺T细胞的比例和活化水平均明显升高，体外抗原刺激可诱导两种品系鸡来源的TCRγδ⁺CD8α⁺ T细胞显著增殖，其中H-B21鸡来源的TCRγδ⁺CD8α⁺ T细胞的增殖启动更早、反应更强。通过实时荧光定量逆转录PCR（RT-qPCR）检测细胞毒性相关基因的表达水平，并采用酶联免疫斑点试验（Enzyme-linked immunospot assay，ELISpot）和胞内细胞因子染色（intracellular cytokine staining，ICS）检测IFN-γ应答，结果表明，活化的TCRγδ⁺CD8α⁺ T细胞具有细胞毒性功能，且H-B21鸡中的应答更为显著。进一步通过分别表达ALV-J的病毒蛋白的细胞作为抗原递呈细胞（Antigen-presenting cell，APC）刺激ALV-J特异性的T细胞，结果显示，Gag和Pol是鸡TCRγδ⁺CD8α⁺ T细胞主要识别的靶抗原。进一步通过ELISpot和ICS分析，发现鸡TCRγδ⁺CD8α⁺ T细胞识别的优势抗原肽为Gag_187-215与Pol_417-445。综上所述，本研究揭示了鸡TCRγδ⁺CD8α⁺ T细胞在ALV-J感染中的应答特征及识别的优势抗原肽，为深入阐明鸡抗ALV-J的细胞免疫调控机制，以及研发基于T细胞表位的新型疫苗提供了重要的理论基础与实验依据。

Abstract

TCRγδ T cells are a major lymphocyte population in chickens, but their response to avian leukosis virus subgroup J (ALV-J) and recognized antigens remain poorly understood. Here, we report an increase in the proportion and activation state of TCRγδ⁺CD8α⁺ T cells in vivo and in ex vivo cultured T cells from 2 chicken lines (MHC homozygous H-B21 lines and outbred G-WL line) with the earlier and stronger proliferative response observed in the H-B21chickens. Activated TCRγδ⁺CD8α⁺ T cells performed ‘cytolytic’ potential via detection of transcription levels of cytotoxic genes with quantitative reverse transcription polymerase chain reaction (qRT-PCR), and IFN-γ protein level with ELISPOT and an intracellular cytokine staining (ICS) assays with more significant response in the H-B21 chickens. Consistently, Furthermore, the viral structural proteins Gag and Pol were identified as major target proteins recognized by chicken TCRγδ⁺CD8α⁺ T cells. Further ELISPOT and ICS analyses mapped the key peptides to Gag_187–215 and Pol_417–445 recognized by Chicken TCRγδ⁺CD8α⁺ T cells against ALV-J infection. Collectively, these findings firstly confirmed that TCRγδ⁺CD8α⁺ T cells could recognize viral antigens and revealed the dominant peptides on ALV-J, enhancing our understanding of anti-ALV-J T cell responses and advancing vaccine design strategies against ALV-J.

Keywords: MHC B21 haplotype chickens ALV-J TCRγδ⁺CD8α⁺T cells chicken T cell culture peptides

Received: 16 March 2026 Accepted: 23 June 2026 Online: 10 July 2026

Fund:

This work was supported by the grants from the National Natural Science Foundation of China (32172868 and 32211530072, to Manman Dai), the Laboratory of Lingnan Modern Agriculture Project, China (NT2025005, to Manman Dai), the Specific University Discipline Construction Project, China (2023B10564003, to Manman Dai), the Young Scholars of Yangtze River Scholar Professor Program, China (2024, to Manman Dai), and the Young Pearl River Scholar of “Guangdong Special Support Plan”, China (2024, to Manman Dai), the Specific University Discipline Construction Project, China (2023B10564003, to Manman Dai).

About author: Xueqing Li, E-mail: lxq996616048@163.com; #Correspondence Manman Dai, E-mail: daimanman1229@scau.edu.cn

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